Mechanisms and Detection of Antimicrobial Resistance

Microorganisms have survived for millions of years because of their ability to adapt to hostile environments. Since the 1940s, bacteria that cause human infections have been exposed to ever-increasing antimicrobial pressure as a result of appropriate and inappropriate use of these agents. In the US, investigators have estimated that >7 million pounds of antimicrobial agents are consumed annually by humans. Approximately 80% of all antibiotics sold (∼29 million pounds) were used in food-producing animals in the US. In 2015, the US Food and Drug Administration (FDA) introduced the Veterinary Feed Directive to promote the judicious use of antibiotics in food-producing animals.

By 2000, despite intensive campaigns aimed at reducing inappropriate use of antimicrobial agents in medicine, investigators estimated that one half of all children in the industrialized world receive an antimicrobial agent annually and that three-fourths of children 1–2 years of age receive these agents. Although antibiotic use from 2000 to 2010 decreased 18% among children and adolescents, the prescription of broad-spectrum antibiotics increased 79%. Even with continued efforts to reduce antimicrobial use in animals and humans, existing levels of antimicrobial resistance are not likely to return to the levels seen before antimicrobial abuse began.

Antimicrobial resistance exacts a high economic cost (∼$60 billion annually in the US). The human cost also is substantial because drug-resistant organisms frequently do not respond to therapy, thus resulting in hospitalization, surgical interventions, increasing use of diagnostic services, and death. Gram-negative bacilli that produce novel β-lactamases with activity against many or, in some cases, all classes of β-lactam agents are emerging at an alarming rate and spreading globally. These include Verona integron–encoded metallo-β-lactamase (VIM), New Delhi metallo-β-lactamase (NDM), and Klebsiella pneumoniae carbapenemase (KPC). An Infectious Disease Society of America initiative to have 10 new systemic drugs available by 2020 against the ESKAPE organisms ( Enterococcus faecium, Staphylococcus aureus, K. pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacteriaceae) has spurred the development and FDA approval of new antibiotics. ^, Although the rate of approved antimicrobial agents is slow compared with previous decades and the challenge of developing new antimicrobial classes remains, at least 39 compounds from different drug classes were in clinical trials in 2014. Beyond the ESKAPE organisms, extensively drug-resistant and totally drug-resistant Mycobacterium tuberculosis is of global concern, especially in locales with high rates of HIV infection.

Genetics of Antimicrobial Resistance

Intrinsic Resistance

Antimicrobial resistance can be attributed to intrinsic cellular properties, intrinsic mutations, acquisition of resistance genes, or intragenic recombination resulting in mosaic genes ( Table 290.1 ). ^, Intrinsic resistance can be caused by inherent properties of micro-organisms, such as cellular membranes, which render them resistant (e.g., gram-negative species to vancomycin). In addition, intrinsic resistance is mediated by the mutation of chromosomal genes. Although acquired resistance mechanisms are more common, many infections are caused by 1 or few species, thus making interspecies gene acquisition difficult in vivo. In these circumstances (as postulated for both P. aeruginosa in the setting of cystic fibrosis and M. tuberculosis ), the organism’s primary antibiotic defense mechanism is intrinsic mutation. ^, Intrinsic mutations that contribute to antimicrobial resistance include variation in the antimicrobial target site (e.g., fluoroquinolones, aminoglycosides), changes in regulatory genes or promoter sequences (e.g., increase in efflux pump or inactivating enzyme expression, decrease in porin expression), and indirect mutations that affect the organism’s mutation rate. ^,

TABLE 290.1

Common Antibacterial Drug Resistance Mechanisms

Drug Class	Resistance Mechanism	Frequency	Examples
Aminoglycosides	Enzymatic inactivation	Common	Phosphotransferases, acetyltransferases, nucleotidyltransferases in Enterobacteriaceae and Enterococcus spp.
	Efflux pump	Uncommon	MexX-MexY efflux pump in Enterobacteriaceae
	Altered binding site	Rare	Streptomycin-resistant Mycobacterium tuberculosis
	Altered uptake	Common	Streptococci and all anaerobic bacteria
β -Lactam agents	Enzymatic inactivation (multiple classes)	Common	Staphylococcus aureus, Haemophilus influenzae, Enterobacteriaceae
	Ambler class A β-lactamase	Common	ESBL-producing: Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis
	Ambler class B β-lactamase	Common	VIM, IMP, and NDM carbapenemases in Acinetobacter baumannii, Pseudomonas aeruginosa, and selected Enterobacteriaceae; chromosomally encoded in Stenotrophomonas maltophilia
	Ambler class C β-lactamase	Common	AmpC chromosomal β-lactamase in Enterobacter spp.
	Ambler class D β-lactamase	Uncommon	Plasmid encoded in P. aeruginosa and selected Enterobacteriaceae
	Altered penicillin-binding proteins	Common	Oxacillin-resistant S. aureus and penicillin-resistant Streptococcus pneumoniae
	Efflux pump	Uncommon	MexAB-OprM pump in P. aeruginosa
	Altered uptake	Uncommon	Loss of OprF and OprD in P. aeruginosa
Colistin	Altered binding	Uncommon	Altered lipopolysaccharide in P. aeruginosa and A. baumannii
Colistin	Efflux pump and capsular overproduction	Rare	K. pneumoniae
Isoniazid	Altered binding	Common	Mutation of katG in M. tuberculosis
Isoniazid	Overexpression of target	Uncommon	Overexpression of inhA in M. tuberculosis
Linezolid	Altered binding	Uncommon	23s rRNA mutations or methyltransferase in Enterococcus spp.
Macrolides and related compounds	Efflux	Common	Expression and alteration of mef pumps in streptococci
Macrolides and related compounds	Altered binding	Common	Methylation of 23S rRNA (erm) conferring resistance to macrolides, streptogramins, and lincosamides in Staphylococcus and Streptococcus spp.
Metronidazole	Inactivating enzyme	Uncommon	Nitroimidazole reductase in Bacteroides spp.
Quinolones	Altered binding	Common	Mutations in gyr conferring resistance in gram-negative organisms; mutations in par conferring resistance in gram-positive organisms
	Efflux	Uncommon	Possible in a variety of gram-negative and gram-positive organisms
	Protective protein	Rare	Protein expressed by K. pneumoniae that binds fluoroquinolones and thus prevents binding to target
Rifampin	Altered binding	Common	Mutations in rpoB gene in S. aureus, M. tuberculosis, and Neisseria meningitides
Tetracyclines	Efflux pump	Common	Gram-positive and gram-negative organisms
Tetracyclines	Protective proteins	Uncommon	Gram-positive and gram-negative organisms
Trimethoprim-sulfamethoxazole	Overproduction of target	Common	Overproduction of DHFR in a variety of bacteria
	Altered binding	Common	Mutated dhfr gene in S. pneumoniae
	Bypass targeted pathway	Uncommon	Occurs in thymidine-dependent S. aureus
Vancomycin	Altered binding	Common	vanA and vanB in Enterococcus
Vancomycin	Overproduction of target	Uncommon	Vancomycin-intermediate S. aureus

AmpC, Ambler class C β-lactamase; DHFR, dihydrofolate reductase; ESBL, extended-spectrum β-lactamase; NDM, New Delhi metallo-β-lactamase; rRNA, ribosomal RNA; VIM, Verona integron–encoded metallo-β-lactamase.

Although bacteria have a low mutation rate (∼10 ⁻⁸ /base pair) to preserve DNA integrity and function, naturally occurring mutations permit evolution toward fitness. Deleterious mutations also can occur, leading to loss of fitness. ^, Induction of a higher mutation rate (i.e., a mutator phenotype) can be beneficial to the organism, particularly when the organism is under selective pressure, such as environmental or antibiotic stress. Fluoroquinolone and aminoglycoside antibiotics can induce a hypermutable state, leading to antimicrobial resistance. ^, The hypermutator state is inducible or transient, and it allows for the survival and expansion of the resistant bacterial population; clinical importance requires further research. ^,

Acquired Resistance

Acquired resistance occurs through horizontal transfer of resistance genes among organisms by conjugation, transformation, or phage-dependent transduction; in some cases, these genes become a stable part of the recipient chromosome. The 2 types of mobile genetic elements are those that move between cells and those that move within a cell. However, elements that move only intracellularly (e.g., gene cassettes, resistance transposons, integrons) can “hitch a ride” on intercellular mobile elements such as plasmids and conjugative transposons.

Plasmids consist of circular, extrachromosomal double-stranded DNA (dsDNA; ∼4–400 kb) that self-replicates. A bacterium can contain multiple compatible plasmids or multiple copies of the same plasmid. Plasmid-based resistance genes can be propagated either by clonal spread of the organism or by horizontal transfer via conjugation. ^, Resistance genes encoding inactivating enzymes for β-lactam agents (including extended-spectrum β-lactamases [ESBLs] and carbapenemases), macrolides, aminoglycosides, and chloramphenicol; efflux genes for macrolides and tetracyclines; and altered targets for sulfonamides have been found on plasmids. Plasmid-mediated resistance genes can also be located on mobile genetic elements that can relocate (either by transposition or site-specific integration) to other plasmids or chromosomes, thereby resulting in the intraspecies and interspecies spread of antimicrobial resistance.

Transposons (∼2–20 kb) contain insertion sequences and a single gene or few linked genes often encoding antimicrobial resistance. In addition, transposons are flanked by inverted sequence repeats and encode for a transposase enzyme required for transposition. Conjugative transposons exist that move directly from one bacterium to another and have been found to mediate the transfer of resistance in gram-positive bacteria. Among the best-studied conjugative transposons are Tn 916 in Enterococcus faecalis encoding tetracycline resistance and Tn 1549 in enterococci encoding vancomycin resistance. Nonconjugative transposons (e.g., resistance transposons) generally are transferred from cell to cell through plasmids and include Tn 1546 implicated in the transfer of vancomycin resistance from Enterococcus to S. aureus .

Integrons include an integrase, mobile gene cassettes, and an integration site for the gene cassettes to allow for site-specific recombination into plasmids or the bacterial chromosome. Integrons themselves are not mobile but facilitate capture of resistance gene cassettes, which subsequently insert into the chromosome. Integrons encoding antimicrobial resistance determinants have been found in certain Enterobacteriaceae organisms, as well as in P. aeruginosa and Acinetobacter species, and they are associated with resistance to β-lactams (including ESBLs), aminoglycosides, chloramphenicol, trimethoprim, and disinfectants. ^, ^, Integrons can contain multiple gene cassettes and therefore often encode for multidrug resistance. Gene cassettes consist of a coding sequence, usually without promoter sequences, followed by an integrase-specific recombination site, and either can exist in a nonfunctional circularized form or are expressed as part of an integron or transposon. More than 100 gene cassettes have been described.

Mosaicism

Intragenic recombination between a sensitive locus on the host bacterial chromosome and related genes from other bacterial species can result in mosaic genes conferring antimicrobial resistance. This recombination-dependent mechanism occurs primarily by direct uptake of naked DNA and is therefore limited to organisms that are naturally transformable. Some mosaic alleles, or polymorphisms, are lost or are present only in low numbers as a result of decreased bacterial fitness. However, if a mosaic allele expresses a phenotype that is favored by antibiotic-selective pressure, the mosaic likely will survive and establish a new, resistant population. Examples of successful mosaicism are the penicillin-binding proteins (PBPs) in Streptococcus pneumoniae and the pathogenic Neisseria species, as well as sulfonamide-resistant dihydropteroate synthase in N. meningitidis. ^,

Mechanisms of Resistance

Bacteria can develop resistance to antimicrobial agents by at least 6 basic mechanisms, and multiple mechanisms can act concurrently:

1.
Enzymatic inactivation
2.
Alteration of the antimicrobial binding site
3.
Active efflux
4.
Alterations in membrane permeability to prevent antimicrobial entry
5.
Alterations in enzymatic pathways so that the targeted enzyme is no longer essential for organism survival
6.
Overproduction of antimicrobial targets

In addition, slow rates of growth as seen in small colony variants or in organisms growing in biofilms also can contribute to resistance in vivo that is not detected readily in vitro when using standard susceptibility test methods.

Aminoglycosides

Antimicrobial resistance in aminoglycosides can be either intrinsic or acquired. Intrinsic resistance is primarily caused by the inability of these molecules to accumulate in the cytoplasm where they must bind to the 30S ribosome to have an antibacterial effect, which is achieved by interfering with binding of transfer RNA (tRNA) to ribosomal RNA (rRNA) during translation and resulting in inhibition of protein synthesis. ^, Aminoglycosides are actively transported into the bacterial cell by a 3-step, energy-dependent process. The energy necessary for this process is generated during aerobic respiration. Bacteria, when growing anaerobically, are not able to generate sufficient energy to “drive” this highly charged molecule into the cell. As a result, all anaerobic organisms and those that depend on anaerobic metabolism (e.g., enterococci) are resistant to aminoglycosides.

Clinically significant, acquired aminoglycoside resistance primarily is the result of acquisition of extrachromosomal elements that encode enzymes that chemically modify aminoglycosides to render them unable to bind to the ribosomal target. Three major classes of enzymes inactivate aminoglycosides, and they are classified by the specific reaction catalyzed: (1) phosphotransferases (APH) that phosphorylate specific aminoglycoside hydroxyl groups, (2) acetyltransferases (AAC) that modify aminoglycosides through acetylation of selected amino groups, and (3) nucleotidyltransferase (ANT) that adenylates the aminoglycoside by adding adenosine monophosphate to selected hydroxyl groups. Resistance in clinical isolates depends on the specific inactivating enzyme produced by the organism. For example, P. aeruginosa can harbor 2 different types of AAC(6′); one type inactivates tobramycin and amikacin, whereas the other inactivates tobramycin and gentamicin. Each enzyme type has variants that can inactivate each of the clinically relevant aminoglycosides. However, certain enzyme types predominate. Because of its modified chemical structure, amikacin is not inactivated by as many of these enzymes, thus resulting in a broader spectrum of activity against gram-negative bacilli compared with gentamicin and tobramycin. Investigators have sought inhibitors of the transferases to allow for increased activity of aminoglycosides in resistant strains, but these studies are preliminary. ^, Aminoglycoside resistance is also important among gram-positive organisms (GPOs). In staphylococci, a unique bifunctional enzyme AAC(6′)-I-APH(2″) produces modification in all three of the most commonly used aminoglycosides (tobramycin, gentamicin, and amikacin), with resulting resistance. Although enterococci are intrinsically resistant to aminoglycosides because of impermeability, exposure to cell wall–active agents such as ampicillin or vancomycin can increase permeability and, in combination with an aminoglycoside, can result in synergistic killing. Resistance secondary to impermeability is referred as “low-level” resistance (gentamicin minimal inhibitory concentrations [MICs], 16–64 μg/mL). High-level resistance also occurs (gentamicin MIC >500 μg/mL), especially among vancomycin-resistant E. faecium strains. Gentamicin resistance in enterococci reflects enzymatic inactivation, primarily the AAC(6′)-I-APH(2″) enzyme. Strains producing this enzyme can be susceptible to high levels of streptomycin.

Aminoglycoside resistance resulting from efflux of aminoglycosides occurs, but it is much less common than enzymatic modification. This efflux pump is a 3-component membrane structure consisting of the proteins MexX-MexY and an outer membrane protein (Opr). Both OprM and OprG act as pores in the outer membrane for these efflux pumps. ^,

Alteration of the antimicrobial binding site as a mechanism of acquired resistance is observed infrequently, and typically it is caused by mutation. Because multiple genes encoding ribosomes are present in most bacteria, the likelihood of having the same random mutational event occurring at the same loci in multiple genes is minimal. The lone exception is in M. tuberculosis. Because it has only a single ribosomal gene copy, ribosomal mutations can confer resistance to streptomycin.

A second means of alteration of the aminoglycoside binding site has been recognized in a strain of K. pneumoniae that possesses a plasmid that encodes a 16S rRNA methyltransferase. Modification of the 16S rRNA results in high-level resistance to gentamicin, tobramycin, and amikacin. Methyltransferase has significant sequence homology with enzymes from aminoglycoside-producing organisms, a finding suggesting that it evolved naturally for self-defense. Ribosomal methylase has been found in transposons within plasmids, thus making horizontal transfer of this resistance gene likely. The 16S rRNA methyltransferase has been detected among Enterobacteriaceae, Acinetobacter, and Pseudomonas .

β-Lactam Agents

Resistance to β-lactam agents is caused primarily by production of β-lactamases or alteration in PBPs. In gram-negative organisms (GNOs), β-lactamase production is the more important resistance mechanism, whereas alteration in PBPs plays a central role in GPOs, including S. aureus, S. pneumoniae, and Enterococcus spp. Organisms can harbor genes simultaneously that encode both resistance mechanisms, as is seen in methicillin-resistant S. aureus (MRSA). Organisms can express >1 β-lactamase gene as well as multiple β-lactam resistance mechanisms concurrently, including β-lactamases, β-lactam efflux, and loss of outer membrane protein porins (important in periplasmic drug accumulation in GNOs).

Classification Systems for β-Lactamases

β-Lactamases are enzymes that degrade the β-lactam ring. They can be encoded chromosomally or on extrachromosomal elements. In GPOs, they are excreted into the extracellular space, whereas in GNOs they are found in the periplasmic space. The Ambler system is a common classification system that classifies the enzyme into 4 different groups (A, B, C, D), based on the enzyme structure. Ambler type A, C, and D β-lactamases are classified as serine β-lactamases because they have serine at the enzyme’s active site. Ambler type B enzyme is classified as a metallo-β-lactamase (MBL) because of the requirement for divalent cations, typically zinc, at the active site.

β-Lactamase-Mediated Resistance

The most common type A β-lactamases are TEM and SHV. These β-lactamases degrade penicillin G, ampicillin, antipseudomonal penicillins, and first-generation cephalosporins. Clavulanic acid has excellent inhibitory activity against these enzymes. Organisms producing these enzymes remain susceptible to aztreonam and third-generation cephalosporins. More than 180 TEM-type enzymes and 130 different SHVs have been identified. TEM frequently is found in Escherichia coli, Haemophilus influenzae, and Neisseria gonorrhoeae, whereas SHV is found in a variety of Enterobacteriaceae members and P. aeruginosa. The most clinically relevant β-lactamase among GPOs is Ambler class A penicillinase-producing S. aureus, rendering resistance to β-lactams. Semisynthetic penicillins such as methicillin, oxacillin, nafcillin, and others were developed that were poorly hydrolyzed by the S. aureus penicillinase, thus maintaining clinical activity.

Extended-Spectrum β-Lactamase Enzymes and Carbapenemase Enzymes

Mutations in the region of the active site of these enzymes can expand their spectra to hydrolyze aztreonam and third-generation cephalosporins such as cefotaxime and ceftazidime. Clavulanic acid continues to inhibit these enzymes in vitro, and these organisms retain susceptibility to cephamycins (cefoxitin) and carbapenems (imipenem, meropenem). Bacteria that are able to express this genotype are described as producing ESBLs. Genes encoding ESBLs frequently are found on large plasmids, which also can encode resistance to other agents such as aminoglycosides and fluoroquinolones. SHV and TEM-type ESBLs are observed most frequently in E. coli, Proteus mirabilis, and K. pneumoniae, and bloodstream infections with these organisms are associated with significantly increased rates of morbidity and mortality.

Another important class A ESBL is CTX-M, so designated because of preferential hydrolytic activity against cefotaxime compared with ceftazidime. More than 90 different types of CTX-M have been identified, and CTX-M has been disseminated on plasmids to many different Enterobacteriaceae organisms, particularly to Salmonella strains originating in South America. As with the ESBL variations of TEM and SHV enzymes, organisms expressing CTX-M are resistant to all classes of penicillin, aztreonam, and first-, second-, and third-generation cephalosporins. Isolates expressing CTX-M remain susceptible to cephamycins and carbapenems, and like other type A ESBLs, this enzyme is inhibited in vitro by clavulanic acid and tazobactam.

The KPC enzymes are an emerging group of class A β-lactamases. Initially associated most closely with K. pneumoniae , plasmids encoding KPC now are found in many Enterobacteriaceae organisms. First identified in 2001, KPCs are now found throughout the US. KPCs are particularly worrisome because they are capable of degrading carbapenems such as imipenem in addition to penicillins, aztreonam, and cephalosporins. KPC-producing strains also can be resistant to aminoglycosides and fluoroquinolones, thus leaving few therapeutic options. Traditional β-lactam–β-lactamase inhibitor combinations do not work in many KPC-expressing strains. However, newer β-lactamase inhibitors such as avibactam and relebactam have activity against KPC-producing organisms and offer newer therapeutic options. ^,

The Ambler class C β-lactamase is referred to as AmpC. AmpC can be encoded chromosomally or on a plasmid. Chromosomally encoded AmpC β-lactamases typically found in Enterobacter and Serratia are inducible and result in low-level resistance to ampicillin, cefazolin, and cefoxitin. Induction by cephamycin or carbapenems can result in high-level resistance to penicillin and first-, second-, and third generation cephalosporins. AmpC is not inhibited by clavulanic acid. Plasmid-encoded AmpC was first reported in K. pneumoniae. Plasmids carrying AmpC often carry resistance genes for a variety of antibiotics, including aminoglycosides, fluoroquinolones, trimethoprim, and tetracylines. Such organisms have constitutive AmpC production and as a result typically are resistant to penicillins, including those used in combination with β-lactamase inhibitors, as well as first-, second- and third-generation cephalosporins. Some, but not all, also are resistant to monobactams. Carbapenems are not well degraded by AmpC and remain a therapeutic option, although isolates with mutation in outer membrane porins can be resistant. Newer agents, specifically ceftolozane and tazobactam, have been developed that retain activity against organisms with AmpC enzymes even with changes in porin permeability or efflux pumps.

The prototypic Ambler class D β-lactamase is the OXA family, so designated because of higher rate of hydrolysis of oxacillin compared with benzylpenicillin. Some OXA enzymes exhibit carbapenemase activity, although this activity may not confer resistance in vivo. Most OXA enzymes are resistant to β-lactamase inhibitors, although avibactam has demonstrated activity against OXA-48. ^, Plasmid-encoded ESBL OXA enzymes have been found in P. aeruginosa . However, the most clinically important OXA-carbapenemase–producing organism are Acinetobacter organisms. In Acinetobacter, OXA-carbapenemases can be encoded chromosomally or on plasmids. Such Acinetobacter organisms are resistant to penicillins, first-, second-, and third-generation cephalosporins, cefepime, and carbapenems. These isolates also are resistant to aminoglycosides and fluoroquinolones. Since the mid-2000s, OXA-carbapenemase–producing Acinetobacter has spread globally and has caused nosocomial outbreaks in numerous facilities.

The Ambler class B β-lactamases are enzymes that require divalent cations, typically zinc, to hydrolyze the β-lactam ring. Class B β-lactamases are capable of hydrolyzing most β-lactam containing antibiotics including carbapenems and are inhibited by chelating agents such as ethylenediaminetetraacetic acid (EDTA) but not by traditional β-lactamase inhibitors. Aztreonam alone among the β-lactams can retain some activity against isolates expressing these enzymes. The class B enzymes can be chromosomally encoded, as is seen with Stenotrophomonas maltophilia, or they can be transferred by plasmids, transposons, or integrons. The most commonly encountered transferable MBLs are IMP and VIM. MBL-producing isolates also frequently are resistant to fluoroquinolones and aminoglycosides. IMP is found most commonly in P. aeruginosa and A. baumannii but also can be found in Enterobacteriaceae, particularly Serratia marcescens, and other Pseudomonas species . VIM is primarily found in P. aeruginosa but has been reported in Enterobacteriaceae and Acinetobacter . The emergence of the NDM is of significant concern. ^, First seen in the UK in 2008, NDMs are currently the most common carbapenemase-producing organisms recovered clinically in the UK. Furthermore, NDMs now have been found in the US, as well as throughout the Pacific region. Importantly, new β-lactam inhibitors such as avibactam or relebactam do not inhibit the activity of MBLs. Treatment requires the use of a combination of alternative agents.

β-Lactam Resistance by Modification of Target Site Binding Proteins

Resistance to penicillinase-stable penicillins developed rapidly in S. aureus as a result of alteration at the specific PBP2 target site. This modified PBP, designated PBP2a, has low affinity for almost all β-lactam antimicrobial agents, with the exception of ceftaroline. PBP2a is primarily encoded by the mecA gene, and S. aureus strains that contain mecA are designated as MRSA. Healthcare-associated MRSA (HA-MRSA) arose and isolates frequently were resistant to other agents, including aminoglycosides, macrolides, and fluoroquinolones. ^, In the late 1990s, community-associated MRSA (CA-MRSA) appeared, differing from HA-MRSA. CA-MRSA strains, although resistant to β-lactams and containing mecA , frequently remain susceptible to other classes of antibiotics. The mecA -encoding region, called the staphylococcal chromosomal cassette (SCC) mec type IVa, in CA-MRSA is much smaller than the SCC most commonly encountered in HA-MRSA and does not contain genes for resistance to other agents. The distinction between HA-MRSA and CA-MRSA has become less clear and less relevant because CA-MRSA–type strains are now found routinely in the healthcare setting. Newer mechanisms of methicillin resistance, such as those encoded by a mecA homologue, mecC , will be important to observe because they may not be detected by current targeted methods ( mecA polymerase chain reaction [PCR], PBP 2a detection).

With the redefining in 2008 of penicillin resistance for S. pneumoniae for nonmeningeal isolates from a penicillin MIC ≥2 to ≥8 μg/mL, and for meningeal isolates to penicillin MIC ≥0.12 μg/mL, restatement of the epidemiology of drug resistance among pneumococci is necessary. Most isolates with penicillin MIC 2–4 μg/mL generally are susceptible to third-generation cephalosporins but frequently are resistant to other classes of antimicrobial agents, including macrolides, sulfonamides, and tetracycline. Isolates with penicillin MIC ≥8.0 μg/mL frequently are resistant to third-generation cephalosporins. Penicillin resistance is caused by alterations in PBP2x, 2b, and 1a that result in reduced binding of β-lactams coupled with increased production of branched-structure muropeptides. ^, Resistance is believed to be encoded by mosaic PBP genes that were transferred to S. pneumoniae from commensal respiratory tract streptococci by transformation and recombination.

Penicillins, specifically ampicillin, are the drugs of choice for treating enterococcal infections, but resistance to cell wall–active agents, β-lactams, and vancomycin is common, especially among isolates of E. faecium. Intrinsically resistant to cephalosporins and aminoglycosides, enterococci have acquired resistance to both ampicillin and vancomycin. β-Lactamase does not play a role in ampicillin resistance of E. faecium . Alteration in PBP5 may be responsible for low-level ampicillin resistance, but assigning it a role in high-level resistance has proven problematic.

β-Lactam resistance among H. influenzae organisms can be caused by β-lactamases or modification in PBP3, so-called β-lactamase–negative ampicillin-resistant (BLNAR) strains. Infrequently, strains are identified that produce β-lactamase and also have altered PBP3. Strains are resistant to ampicillin–β-lactamase inhibitor agents. ^, β-Lactamases of H. influenzae are Ambler type A, and the encoding genes are found on plasmids. A survey of 6642 H. influenzae isolates collected globally identified 25% producing β-lactamase and 1% with altered PBP. However, certain geographic “hot spots” (e.g., Japan and Spain) have higher rates of resistance because of PBPs. ^,

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